The first tests for detecting pathologies or problems associated with the ingestion of fats involved the obtention of stool samples and the measurement of their fat content. This method had associated with it the problems of collecting and analyzing stools.
Partially in response to these problems, alternative techniques for detecting problems with the ingestion or metabolism of fats using isotope-labeled fats were developed, as disclosed by Caspary, Clin. Gastroenterol 7:351-374 (1978). In this type of test, carbon 14 labeled triolein is administered orally to patients and the amount of carbon 14 (.sup.14 CO.sub.2) radioactivity in the breath is measured. The amount of radioactivity provides a means of diagnosing fat malabsorption. The rate of appearance of the labeled CO.sub.2 in the breath can be used to indicate various pathologies associated with the malabsorption of the substrate. In other words, the rate of appearance will indicate GI tract dysfunction. Also, the rate can be used to detect problems associated with the metabolism of the substrate, for example, liver or pancreas disorders. Additionally, the complete absence of labeled CO.sub.2 in the breath is indicative of metabolic problems. Concerns over the long term effects of the ingestion of radioactive substances have precluded this test from being used with children and with women of childbearing age.
Watkins, et al., Gastroenterology 82:911-7 (1982), disclosed the diagnosis and differentiation of fat malabsorption using .sup.13 C-labeled lipids in breath tests. In general, a .sup.13 C breath test involves the ingestion by the patient of a labeled substrate, i.e. fatty acid, labeled with .sup.13 C, followed by monitoring the carbon dioxide ("CO.sub.2 ") in the exhaled breath of the patient using isotope ratio mass spectrometer ("IRMS") for the appearance of the labeled carbon over a period of time. Normal metabolism produces a rapid appearance of .sup.13 C in the exhaled carbon dioxide as the fatty acids are metabolized. Useful labeled lipids included trioctanoin, triolein and palmitic acid. Triolein has been found to be a preferred substrate.
Triolein is a chemically-synthesized oil comprised of three oleic acid moieties esterified to glycerol, forming a typical vegetable oil-like triglyceride.In the past, the oleic acid moieties have been labeled with either .sup.14 C or .sup.13 C at only the C1 position, in part due to the widespread belief that not all carbons along the fatty acid chain are oxidized equivalently. Also, organic labeling reactions are simpler when only C1 is labeled. Since carbons other than C1 in the fatty acid were thought not to be equivalent to C1 in the fatty acid oxidation pathway only C1 was labeled.
Commercially available .sup.13 C-labeled triolein is a chemically synthesized product, which heretofore has been quite costly to produce. Typically, the selling price is between $700--$1,000 per gram. Inasmuch as a dose of 3-4 grams of lipid or about 50 mg/kg body weight is required to perform a proper diagnostic breath test in adult males, the test is a very costly one to conduct.
Replacement of the radioactive carbon with its stable isotope and subsequent detection using an isotope ratio mass spectrometer has now made the breath test safe for any population of subjects. However, because of the prohibitive costs associated with chemically synthesizing labeled triolein, the test has not yet enjoyed widespread success.
Accordingly, it would be desirable to have a method of producing labeled compounds for use in diagnostic breath tests which significantly reduces the cost of such compounds.
It is a further object of this invention to provide a method for obtaining fatty acids and triglycerides with high levels of .sup.13 C-enrichment for use in diagnostic tests. Such compounds would increase the sensitivity of the test and further reduce the cost.